Exploration of urine and plasma biomarkers in liver fibrosis and hepatocellular carcinoma

Abstract

Liver fibrosis is a major risk factor for development of hepatocellular carcinoma. Both liver fibrosis and hepatocellular carcinoma are associated with molecular pathogenic mechanisms involving alterations in the hepatocellular proteome, metabolome, and genome. Both liver fibrosis and hepatocellular carcinoma lack suitable biological predictive biomarkers in clinical practice. Therefore, to aid in identifying suitable biomarkers, three approaches were employed in patients with liver fibrosis and hepatocellular carcinoma. Firstly, proteomic analysis was applied to identify post-translational enzymatic protein modifications peripherally present in the urine. Secondly, metabolic profiling was applied to characterise small volatile organic compounds present in the urine. Thirdly, DNA methylation detection technology was applied, to identify methylated SEPTIN9 patterns among the circulating hepatocellular carcinoma DNA molecules within the cell-free DNA pool present peripherally in the plasma. Urinary proteomic analysis identified novel specific peptides for liver fibrosis and hepatocellular carcinoma. Additionally, proteases potentially involved in liver fibrosis and hepatocellular carcinoma were predicted from the peptides sequence with further demonstration of these proteases by immunohistochemistry in human normal liver tissue, liver fibrosis and hepatocellular carcinoma. The identified urinary peptides showed good diagnostic and prognostic performance in liver fibrosis and hepatocellular carcinoma. Urinary metabolic profiling technologies demonstrated that volatile organic compounds patterns can be used noninvasively to detect hepatocellular carcinoma and they also revealed chemical composition of novel volatile organic compounds related to liver fibrosis and hepatocellular carcinoma. DNA methylation analysis showed that methylated SEPTIN9 has good sensitivity and specificity for hepatocellular carcinoma. It was also a prognostic indicator in patients with liver disease and hepatocellular carcinoma. The methylated SEPTIN9 was also associated with other surrogate biomarkers for liver function, liver fibrosis and inflammation. Additionally, methylated SEPTIN9 was noted to incrementally increase in various stages of liver disease. The researched biomarkers in this work provided some insight into the pathogenic mechanisms of liver fibrosis and hepatocellular carcinoma. If further validated, the identified biomarkers in this work could offer cost-effective tools for screening, diagnosis, prognosis and/or surveillance, particularly in low resource settings where access to advanced imaging and invasive biopsy is not feasible